I am trying to amplify the hypervariable polymorphism in the APOC3 gene, which was described by Bhattacharya et al. (NAR 1991 vol 19:4799. I was successful at the first attempt. I am now completely unable to reproduce this amplification, although PCR conditions are the same as the first time. This is really puzzling.
This polymorphism is 350 bp long and rich in GC. I am using 0.5 ng per microliter of each primer, 1 U of taq, 1.4 mmol MgCl2, and 1/100 vol PerfectMatch.
Do you have encountered this kind of problem? Can you help?
Thanks. q